The experiments outlined in this revised proposal reflect the PI's longstanding interest in the role of amyloidotic and neurofibrillary lesions in the pathogenesis of Alzheimer's disease (AD). They focus on early events which may precede the formation of these pathological abnormalities. Because there is yet no evidence that the Beta-amyloid protein precursor (betaAPP or pre-A4) gene is abnormal in AD, these studies concentrate on possible transcriptional or posttranslational events which could explain the excess progressive deposition of beta-amyloid protein (betaAP or A4) in the brain of patients with AD, as compared to aged normals. It is unlikely that a single abnormality at the protein level will account for the complex spectrum of pathological lesions in AD. For example, the extent of vascular amyloidosis varies widely among AD brains, even among individuals with comparable numbers of plaques. Also, many investigators now agree that there is a form of AD which lacks neocortical tangles, and that this subset of patients cannot be distinguished from AD cases with abundant tangles by clinical or biochemical criteria. Hence, the proposed experiments address a series of interrelated questions aimed at several aspects of altered brain structure and function in AD. Specific Aim #1 analyzes what may be the most prevalent lesion in AD brains - diffuse ("pre-amyloid") plaques. These lesions have been defined as amorphous betaAP deposits apparently lacking reactive neurites or glial cells. We hypothesize that diffuse plaques may be a very early lesion and one that contains betaAP in a non-fibrillar (i.e., not amyloid) form. Specific Aim #2 explores experiments aim to confirm and expand our strong preliminary data showing specific betaAP immunostaining in AD and aged normal skin and colon, but not tissues from young (<60 yrs) controls. Confirmation of this new data carries implications about the basic mechanisms of betaAP deposition. Specific Aim #3 proposes a detailed EM study of amyloidotic vessels to look for the earliest immunocytochemically detectable betaAP deposits and asks if betaAP is present in vessels in a non-fibrillar form. In addition, the experiments will determine if the vascular distribution of beta-amyloid, which is known to be segmental and focal, can be correlated with that of smooth muscle cells and/or pericytes. If so, these cells may be important in the processing/deposition of betaAP in vessels. Specific Aim #4 addresses several issues concerning the normal function of betaAPP, using human cell lines stably transfected with full length betaAPP cDNAs. The emerging theme of growth-promoting properties of betaAPP will be examined. Taken together, the proposed experiments address features of AD which may represent important pathogenetic steps in the evolution of this common disease. This FIRST Award represents a critical step in the dedication of the PI's career to research on human neurodegenerative disease.